Abstract

Cellular senescence, a stress-induced irreversible growth arrest often characterized by expression of p16(Ink4a) (encoded by the Ink4a/Arf locus, also known as Cdkn2a) and a distinctive secretory phenotype, prevents the proliferation of preneoplastic cells and has beneficial roles in tissue remodelling during embryogenesis and wound healing. Senescent cells accumulate in various tissues and organs over time, and have been speculated to have a role in ageing. To explore the physiological relevance and consequences of naturally occurring senescent cells, here we use a previously established transgene, INK-ATTAC, to induce apoptosis in p16(Ink4a)-expressing cells of wild-type mice by injection of AP20187 twice a week starting at one year of age. We show that compared to vehicle alone, AP20187 treatment extended median lifespan in both male and female mice of two distinct genetic backgrounds. The clearance of p16(Ink4a)-positive cells delayed tumorigenesis and attenuated age-related deterioration of several organs without apparent side effects, including kidney, heart and fat, where clearance preserved the functionality of glomeruli, cardio-protective KATP channels and adipocytes, respectively. Thus, p16(Ink4a)-positive cells that accumulate during adulthood negatively influence lifespan and promote age-dependent changes in several organs, and their therapeutic removal may be an attractive approach to extend healthy lifespan.

a, Closure of 3-mm punch biopsy wounds in 18-month-old ATTAC females after treatment with vehicle or AP for 6 months and if drug treatment was stopped 2 days prior to skin puncture or continued during wound closure (n = 6 wounds for –AP;–AP and +AP;–AP and n = 10 wounds for –AP;+AP and +AP;+AP). AP administration during the wound healing process significantly attenuates the rate of wound closure independently of whether senescent cell removal had occurred prior to wounding. b, Closure of 3-mm punch biopsy wounds in 4-month-old ATTAC females after treatment with vehicle or AP following wounding (n = 10 wounds per group). Similar to 18-month-old mice, AP administration during the wound healing process dramatically attenuated the rate of wound closure. c, Quantification of total GFP+ cells isolated from 3-mm punch biopsy wounds of 4-month-old mice two days into the wound healing process treated with vehicle (black) or AP (red, n = 3 mice per group). d, PTAH-stained tissues sections from 18-month-old ATTAC mice for detection of fibrosis. Scale bars, 100 μm. Error bars indicate s.e.m. Unpaired two-tailed t tests were used to determine statistical significance for a–c. Mice receiving AP during the healing process in a and b are significantly different from those treated with vehicle from day 1.5 through day 9.5. *, P<0.05.

a, Study design for clearance of senescent cells in mixed and C57BL/6 mouse cohorts. Healthspan analysis was done at 18 months, an age where relatively few mice in vehicle (Veh)- or AP-treated have died and bias due to selection for long-lived animals is unlikely. b, c, Survival curves for vehicle- (–AP) and AP-treated (+AP) mixed (b) and C57BL/6 mice (c). Median survival (in days) and percentage increase in median survival are indicated. We note that median lifespans of our vehicle-treated cohorts are similar to those of wild-type mice administered AP (c). Log-rank tests were used to determine statistical significance. *, P<0.05; **, P<0.01; ***, P<0.001.

a, Survival curves of AT-TAC mice dying of cancer (mice that had an overt tumor at time of death; mice with lymphomas, sarcomas and carcinomas were included, mice without tumors were censored). Median survival and percentage increase are indicated. b, Representative images of aged mice with and without senescent cell clearance. c, Spontaneous movement and exploratory behavior of ATTAC mice analyzed by the open field test. Error bars indicate s.e.m. Log-rank tests were used to determine statistical significance in a; unpaired two-tailed t tests were used in c. *, P<0.05; **, P<0.01; ***, P<0.001.